PROJECT1:SYSTEMATICIDENTIFICATIONOFDRIVERNETWORKSINCANCER SUMMARY Avastnumberofmutationscontributetocancer,buttheobservednonrandomcombinationsofthoseleading to transformation highlight the importance of hallmark pathways and networks in cancer progression. While manypathwayshavebeenimplicatedincancer,attributessuchastumorheterogeneity,tissueoforigin,and degreeofprogressionleadtoeachcaseexhibitingauniquesubsetofalteredpathways.Takentogether,this diversity among cancer types and their origins has complicated the development of targeted cancer treatments.Weproposeheretosystematicallyidentifytheproteinnetworksthatdrivecancer,acrossarange of tumor types starting with head and neck squamous cell carcinoma (HNSCC) and breast cancer (BC). Coupled with functional validation and highresolution structural analysis of the key protein interactions and complexes,weanticipatemajorinsightsintotheunderlyingtumorbiologyaswellasthepotentialtounravel geneticvulnerabilitiesoftherapeuticrelevance. In?Project1?,CCMIinvestigatorswillbuildaphysicalinteractionmappingpipelinefocusedonunderstanding the underlying network biology behind cancer. To this end, we are targeting 80 genes genetically linked to eitherHNSCCorBCandsubjectingthewildtypeproteinsandnumerousmutantformstoaffinitypurification massspectrometry(APMS)inapanelofrelevantcancersubtypecelllines(?Aim1?).Tocomplementthese datawewillperformfunctionalkinomescreensusingthehighthroughputkinaseactivitymapping(HTKAM) platform,whichwillquantifyhowkinasesignalingnetworksarerewiredbydifferentproteinmutations,andin differentcellularbackgrounds.Next,wewillusethecomputationaltechniqueofnetworkpropagationtodefine themajormutateddriverpathwaysunderlyingeachdiseasesubtype,inwhichphysicalproteininteractionsare integrated with somatic and germline mutations identified in tumor genomes. The results of this network characterization(?Aim1?)andintegrativeanalysis(?Aim2?)willidentifynetworkcomponentsthatcouldserveas targetsfortherapeuticintervention?in?Aim3wewillperformcellularassaystovalidatethesenetworktargets. Lastly, in ?Aim 4 ?we will use cryogenic electron microscopy (cryoEM) to structurally characterize therapeuticallyactionableproteincomplexesanddeveloptechnologytoenablethescreeningofmanymore. Successfulcompletionofthisworkwillyieldanetworkmappingpipelinethatcanbeextendedtomanycancer typesandwillaidintherationalselectionoftherapeutictargetswithgreaterprecisionandspeed.